This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. During large dsDNA virus assembly, viral DNA is transferred into preformed protein shells. The DNA packaging is driven into the shell by a translocation motor system powered by ATP hydrolysis. A crucial part of the packaging machine is a portal protein. To date there is limited detail structural information of portals as only two portal phages SPP1 and phi*29 phage portals have been determined. We have determined the first crystal structure of the double- stranded DNA bacteriophage HK97-like connector to 2.9 A resolution. This 400 kDa motor protein connects the head of the phage to its tail and translocates the DNA into the prohead during packaging. Each monomer has an elongated shape and is composed of a central, alpha-helical domain that includes a distal alpha/beta domain and a proximal six-stranded SH3-like domain. The protomers assemble into a 12-mer, funnel-like structure with a 40 A wide central channel. The surface of the channel is mainly electronegative, but it includes three positively charged rings, one at the opening of the funnel, second at the middle of a particle and the third at the exit of the portal. We are looking to combine the crystal structure with macromolecular imaging in order to extensively characterize this portal structure.